Screening the biosphere: the fungicolous fungus Trichoderma phellinicola, a prolific source of hypophellins, new 17-, 18-, 19-, and 20-residue peptaibiotics.

To investigate the significance of antibiotics for the producing organism(s) in the natural habitat, we screened a specimen of the fungicolous fungus Trichoderma phellinicola (syn. Hypocrea phellinicola) growing on its natural host Phellinus ferruginosus. Results revealed that a particular group of non-ribosomal antibiotic polypeptides, peptaibiotics, which contain the non-proteinogenic marker amino acid, α-aminoisobutyric acid, was biosynthesized in the natural habitat by the fungicolous producer and, consequently, released into the host. By means of liquid chromatography coupled to electrospray high-resolution time-of-flight mass spectrometry, we detected ten 20-residue peptaibols in the specimen. Sequences of peptaibiotics found in vivo were independently confirmed by analyzing the peptaibiome of an agar plate culture of T. phellinicola CBS 119283 (ex-type) grown under laboratory conditions. Notably, this strain could be identified as a potent producer of 39 new 17-, 18-, and 19-residue peptaibiotics, which display the same building scheme as the 20-residue peptaibols found in the specimen. Two of the 19-residue peptaibols are tentatively assigned to carry tyrosinol, a novel C-terminal residue, as deduced from high-resolution tandem mass-spectrometry data. For the new peptaibiotics produced by T. phellinicola, the name 'hypophellin(s)', based on the teleomorph name, is introduced.

The sequences of the eleven-residue peptaibiotics: suzukacillins-B.

The filamentous fungus designated 'Trichoderma viride' strain 63 C-1 simultaneously produces suzukacillins (SZs), two microheterogeneous groups of peptaibols, under submerged culture conditions. Both groups are readily distinguishable by TLC: the major group is designated SZ-A, whereas the minor group with a higher Rf value is named SZ-B. The peptide mixture was obtained from a MeOH extract of the mycelium. SZ-B was separated from SZ-A by Sephadex LH-20 column chromatography. Although it provided one single spot on silica-gel TLC plates, 15 individual peptides could be separated by C8 reversed-phase (RP) HPLC, and their sequences were determined by HPLC/QqTOF-ESI-HRMS. Fourteen peptides exhibit the C-terminal sequence Pro(6) -Lxx-Lxx-Aib-Pro-Vxxol/Lxxol(11) , which is common for eleven-residue peptaibols. The remaining peptide is tentatively assigned as a ten-residue sequence, in which the C-terminal 1,2-amino alcohol is deleted, thus terminating in free proline. Nine of the peptides carry an Ac-Aib residue at the N-terminus, very frequently found in eleven-residue peptaibols. Four peptides comprise the rare Ac-Ala N-terminus, and for two peptides, N-terminal Ac-D-Iva residues were identified. One peptide contains a C-terminal residue of yet undetermined structure. Comparison with previously reported eleven-residue peptaibol sequences reveals that eight of the peptides represent new sequence analogs.

Hypopulvins, novel peptaibiotics from the polyporicolous fungus Hypocrea pulvinata, are produced during infection of its natural hosts.

In order to investigate the significance of antibiotics for the producing organism(s) in the natural habitat, we screened specimens of the polyporicolous fungus Hypocrea pulvinata growing on its natural hosts Piptoporus betulinus and Fomitopsis pinicola. Results showed that a particular group of nonribosomally biosynthesised antibiotic polypeptides, the peptaibiotics, which contain the nonproteinogenic marker amino acid α-aminoisobutyric acid (Aib), was produced in the natural habitat by the fungicolous producer and, consequently, released into the host. Using liquid chromatography coupled to electrospray high-resolution mass spectrometry we detected especially 19-, but also 11-, 18-, and 20-residue peptaibiotics in the five infected specimens analysed. Structures of peptaibiotics found were confirmed by analysing the peptaibiome of pure agar cultures obtained by single-ascospore isolation from the specimens. The 19-residue peptaibols were determined as deletion sequences of the trichosporins B lacking the Aib residue in position 6. Notably, 26 of the 28 peptaibiotics sequenced were novel; therefore the name 'hypopulvins' was introduced. Considering not only the ubiquity of both the two host species but also the highly specific association between H. pulvinata and P. betulinus/F. pinicola, and the abundance of this fungicolous species in north temperate regions of the world, a decisive role for the peptaibiotics detected in this study is predicted, which may act as mediators of the complex interactions between the basidiomycetous host and its fungicolous ascomycete 'partner'. Structural analogies of the hypopulvins, particularly with other 18-, 19-, and 20-residue peptaibiotics, suggest that the hypopulvins are forming transmembrane ion channels and could thus support the hypothesis of a parasitic lifestyle of the fungicolous producer.

Harmonine, a defence compound from the harlequin ladybird, inhibits mycobacterial growth and demonstrates multi-stage antimalarial activity.

The harlequin ladybird beetle Harmonia axyridis has been introduced in many countries as a biological control agent, but has become an invasive species threatening the biodiversity of native ladybirds. Its invasive success has been attributed to its vigorous resistance against diverse pathogens. This study demonstrates that harmonine ((17R,9Z)-1,17-diaminooctadec-9-ene), which is present in H. axyridis haemolymph, displays broad-spectrum antimicrobial activity that includes human pathogens. Antibacterial activity is most pronounced against fast-growing mycobacteria and Mycobacterium tuberculosis, and the growth of both chloroquine-sensitive and -resistant Plasmodium falciparum strains is inhibited. Harmonine displays gametocytocidal activity, and inhibits the exflagellation of microgametocytes and zygote formation. In an Anopheles stephensi mosquito feeding model, harmonine displays transmission-blocking activity.